The superfamily of G protein-coupled receptors (GPCRs) comprises plasma membrane spanning proteins that transduce signals via heterotrimeric G proteins on the inner surface of the plasma membrane, leading to intracellular signaling cascades. Jacoby et al. (2006) ChemMedChem 1:761-782. The cell surface location, tissue distribution, and diversity of these GPCRs make them ideal targets for drug intervention. It is widely reported that roughly 30% of marketed drugs target specific GPCR activity. Jacoby, E. B. (2006), supra; Overington et al. (2006) Nature Reviews Drug. Disc. 5:993-996.
The Opioid Receptors (ORs) are members of the Class A family of GPCRs. As such, ORs mediate the actions of endogenous opioids (e.g., endorphins) as well as the action of exogenous opioids such as morphine and morphine-like opiates, including most clinical analgesics. Four OR types are known to exist: the mu OR, the delta OR, the kappa OR and the L1OR; respectively abbreviated as MOR, DOR, KOR, ORL1. These OR subtypes appear to have overlapping functional mechanisms at a cellular level, share roughly 60% amino acid identity, and signal through the Gi/o family of heterotrimeric G proteins. The signaling pathways of opioid receptors are well characterized. Stein & Machelska (2011) Pharmacological Reviews 63:860-881.
After binding of an orthosteric ligand, conformational changes in the receptor allow intracellular coupling of heterotrimeric Gi/o proteins to the C terminus of the receptor. At the Gα subunit, GTP replaces GDP and dissociation of the trimeric G protein complex into Gα and Gβγ subunits ensues. Subsequently, these subunits can inhibit adenylyl cyclases and thereby reduce cAMP production and/or directly interact with different ion channels in the membrane.
The different OR types appear to share many functional mechanisms at the cellular level. The activation and/or inactivation of the ORs by endogenous or exogenous orthosteric opioids results in inhibition of adenylyl cyclase, modulation of ion channel activity, and transcriptional changes in the cell. Waldhoer et al. (2004) Annual Review of Biochemistry 73:953-990. For example, activation of the mu opioid receptors causes inhibition of adenylate cyclase (resulting in lower intracellular cAMP levels), and recruits β-arrestin to the receptor. β-arrestin recruitment is a non-G-protein mediated signaling pathway through which many GPCRs (including the ORs) signal. Evidence exists that β-arrestin is involved in receptor desensitization and internalization/recycling. Whalen, E. R. (2011) Trends in Molecular Medicine 17:126-139; Shukla et al. (2011) Trends in Biomedical Sciences 36:457-469.
ORs are key targets in the management of pain, with morphine and its derivatives inducing pain relief by acting as full or partial receptor agonists. Pain relief (analgesia) is attributed to the actions of opiates and opioids specifically at MOR. Matthes et al. (1996) Nature 383:819-823; Manglik et al. (2012) Nature 485:321-326. When therapeutic doses of morphine are given to patients with pain, the patients report that the pain is less intense, less discomforting, or entirely absent. However, in addition to providing relief of distress, the narrow therapeutic window for morphine also clinically manifests a variety of adverse side effects. Moreover, when a pain-relieving dose of morphine is administered to a pain-free individual, the experience is not always pleasant; nausea is common, and vomiting may also occur. In addition, drowsiness, inability to concentrate, difficulty in mentation, apathy, lessened physical activity, reduced visual acuity, and lethargy may ensue.
Two highly selective MOR agonists, endomorphin-1 (EM1) and endomorphin-2 (EM2), have been isolated from bovine as well as human brains in large quantities and are believed to be the endogenous ligands for the MOR. Zadina et al. (1997) Nature 386:499-502; Hackler et al. (1993) Neuropeptides 24:159-164; Erchegyi et al. (1992) Peptides 13:623-631. Relative to other orthosteric agonists, EM1 and EM2 display an exceptionally high level of binding affinity and selectivity for MOR over KOR and DOR. Zadina et al. (1994) Life Sciences 55:461-466. The differential distribution of these peptides in various tissues has been widely studied with results indicating that EM1 is more densely distributed throughout the brain; whereas EM2 is more prevalent in the spinal cord. Martin-Schild et al. (1999) J Comp. Neurol. 405:450-471. Additionally, the presence of the endomorphins (EMs) and MOR has been confirmed in animal and human models of inflammatory and neuropathic pain. Troung et al. (2003) Ann. Neurol. 53:366-375; Straub et al. (2008) Arthritis and Rheumatism 58:456-466; Yang et al. (2014) PLOS ONE 9(2):e89583; Stein et al. (1993) Lancet 342:321-324; Mousa et al. (2002) J. Neuroimmunol. 126:5-15; McDougall et al. (2004) Am. J. Physiol. Regul. Integr. Comp. Physiol. 286:R634-R641; Obara et al. (2004) Neuroscience Letters 360:85-89. The antinociceptive actions of exogenously administered EM1 and EM2 have been studied in a variety of animal models, as well as in humans. Soigner et al. (2000) Life Sciences 67:907-912; Horvath et al. (1999) Life Sciences 65:2635-2641; Macdougall et al. (2004) J. Molec. Neurosci. 22:125-137; Horvath (2000) Pharmacology & Therapeutics 88:437-463; Przewlocka et al. (1999a) Eur. J Pharmacol. 367:189-196; Przewlocka et al. (1999b) Ann NY Acad. Sci. 897:154-164.
Opioid receptors have been extensively studied because of the needs for (1) better pain control and (2) reduction or elimination of adverse side effects. The side effects common to orthosteric ligands for the opioid receptors include, in addition to those mentioned above, tolerance, respiratory suppression, constipation, allodynia, and dependence. Waldhoer et al., supra; McNicol et al. (2003) J. Pain 4:231-256. Indeed, recent determinations of the therapeutic indices for commonly used opioids led to the conclusion that systemic side effects are to be expected for all of them (Kuo et al. (2015) British J. Pharmacol. 172:532-548), leading some to conclude that the side effects are mechanism-based. Alternatively, the side effects of opioid use could be attributed to the signal bias that these orthosteric ligands induce and/or to the presence of receptors and/or their ligands in tissues that are not experiencing pain. Kenakin (2015a) Trends Pharmacol. Sci. 36:705-706; Stein & Machelska, supra.
To overcome the side effects associated with traditional OR orthosteric agonists and partial agonists, drug discovery efforts have focused on (1) developing selective orthosteric ligands which display OR subtype selectivity either as full agonists, partial agonists or when used in combination therapy (Davis (2012) Exp. Opin. Drug Discov. 7:165-178; Dietis (2009) Br. J. Anaesth. 103:38-49); (2) physiological compartmentalization of orthosteric ligands (Spahn et al. (2017) Science 355:966-969); and (3) inducing selective signal bias of orthosteric ligands (Soergel et al. (2014) Pain 155:1829-1835; Chen et al. (2013) J. Medicinal Chem. 56:8019-8031). These diverse approaches have in common a strategy in which the orthosteric ligand binding domain is the only entity being probed and/or modified. Given the current national epidemic of orthosteric opioid agonist abuse, a novel approach to pain mitigation would provide a welcome benefit to society. Lauren & Rossen (2016) Drug Poisoning Mortality: United States, 2002-2014. Atlanta: National Center for Health Statistics, Centers for Disease Control and Prevention.
Allosteric modulators of opioid receptors provide an alternative strategy for pain mitigation. An allosteric modulator has no intrinsic agonist or antagonist activity toward a receptor but, in the presence of an orthosteric agonist, can further increase the activity of the receptor beyond that induced by the orthosteric agonist (a positive allosteric modulator, or PAM) or decrease receptor activity below that which would normally be induced by the orthosteric agonist (a negative allosteric modulator, or NAM). Thus, the use of positive allosteric modulators of the μ-opioid receptor (MOR PAMS or MOR-PAMs), when used in conjunction with exogenous opioids, would allow lower doses of opioid to be administered, thereby lessening side effects and potential for abuse.
In addition, because endogenous MOR agonists are not distributed throughout the entire body, but tend to be released in areas experiencing pain, the use of a MOR PAM as an analgesic (in the absence of an exogenous opioid) also has the advantage that the body of the subject is not being flooded with an exogenous opioid. Rather, the MOR-PAM activates the receptor only in the regions of the body that already contain endogenous receptor agonists; thereby potentiating the activity of the agonist and activating the receptor only in the regions where necessary for pain reduction.
The naturally-occurring hallucinogen Salvinorin A has been observed to behave, in vitro, as an allosteric modulator of the MOR. Rothman (2007) J. Pharmacol. Exp. Therapeutics 320:801-810. However, further use of this compound for in vivo mechanistic testing and/or therapeutic use is unlikely due to its inherent polypharmacology.
Thiazolidine-based allosteric modulators of the MOR have also been described and tested in vitro. WO 2014/107344; Burford et al. (2011) Biochemical Pharmacol. 81:691-702; Burford et al. (2013) Proc. Natl. Acad. Sci. USA 110:10830-10835; Burford et al. (2015) Br. J. Pharmacol. 172:277-286; Livingston et al. (2014) Proc. Natl. Acad. Sci. USA 111:18369-18374. However, in vitro analyses of these compounds have revealed that they are unlikely to be useful for in vivo studies and routine therapeutic use, because of the difficulty of their synthesis, poor potency and metabolic instability. Some of these compounds also possess pure agonist activity in addition to PAM activity in certain assays (i.e., they act as “ago-PAMs”) and therefore lack the benefit of being able to amplify endogenous analgesic mechanisms in a temporally- and spatially-limited fashion. (Bisignano et al. (2015) J. Chem. Inf. Model 55:1836-1843; Rockwell & Alt (2017) “Positive Allosteric Modulators of Opioid Receptors” in RSC Drug Discovery Series No. 56: Allosterism in Drug Discovery, ed. D. Doller, Royal Society of Chemistry, Chapter 9, pp. 194-219; Livingston et al., supra.
Modification of these thiazolidine-based MOR PAMs, guided by structure/activity relationship (SAR)-based lead optimization, and attendant pharmacophore modeling, have been conducted. (Bisignano et al. (2015) J. Chem. Inf. Model 55:1836-1843; Bartuzi et al. (2016) J. Chem. Inf. Model 56:563-570. These efforts have also not yielded potent or selective compounds viable for in vivo testing. In addition, the SAR-based derivatives are somewhat non-specific, showing some PAM activity at the delta opioid receptor as well. Rockwell & Alt, supra.
For all of the foregoing reasons, there is a need for new compounds that induce OR signaling and analgesia, but have reduced side effects, and that are suitable for in vivo testing and administration. There is a continuing need for new analgesics that can incorporate the profound beneficial effects of opioids without the concomitant side effects; and for compounds that can potentiate the temporally- and spatially-restricted activity of endogenous opioids.